Materials and Methods of Producing 7-Carbon Monomers

Abstract

This document describes materials and methods for producing 7-hydroxyheptanoic acid using a β-ketothiolase or a synthase and an alcohol O-acetyltransferase to form a 7-acetyloxy-3-oxoheptanoyl-CoA intermediate. This document describes biochemical pathways for producing 7-hydroxyheptanoic acid using a β-ketothiolase or a synthase and an alcohol O-acetyltransferase to form a 7-acetyloxy-3-oxoheptanoyl-CoA intermediate. 7-hydroxyheptanoic acid can be enzymatically converted to pimelic acid, 7-aminoheptanoic acid, heptamethylenediamine or 1,7 heptanediol. This document also describes recombinant hosts producing 7-hydroxyheptanoic acid as well as pimelic acid, 7-aminoheptanoic acid, heptamethylenediamine and 1,7 heptanediol.

1 . A method of producing 7-acetyloxy-3-oxoheptanoyl-CoA, said method comprising enzymatically converting 5-acetyloxypentanoyl-CoA to 7-acetyloxy-3-oxoheptanoyl-CoA using a polypeptide having the activity of a β-ketothiolase or synthase classified under EC. 2.3.1.-. 
     
     
         2 . The method of  claim 1 , wherein said polypeptide having the activity of a β-ketothiolase or synthase is classified under EC 2.3.1.16, EC 2.3.1.41, EC 2.3.1.174, EC 2.3.1.179 or EC 2.3.1.180. 
     
     
         3 . The method of  claim 1 , wherein said polypeptide having the activity of a β-ketothiolase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 1 or 16 or is any other polypeptide having the activity of a β-ketothiolase classified under EC 2.3.1.16 or EC 2.3.1.174. 
     
     
         4 . (canceled) 
     
     
         5 . The method of  claim 1 , further comprising enzymatically converting 7-acetyloxy-3-oxoheptanoyl-CoA to 7-hydroxyheptanoate using a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, a polypeptide having the activity of an enoyl-CoA hydratase, a trans-2-enoyl-CoA reductase, an esterase, and a polypeptide having the activity of a thioesterase or a CoA transferase. 
     
     
         6 . The method of  claim 5 , wherein:
 (a) said polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase or said 3-oxoacyl-CoA reductase is classified under EC 1.1.1.35, EC 1.1.1.36, EC 1.1.1.100, or EC 1.1.1.157;   (b) said polypeptide having the activity of an enoyl-CoA hydratase is classified under EC 4.2.1.17 or EC 4.2.1.119;   (c) said polypeptide having the activity of a trans-2-enoyl-CoA reductase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 23-28 or is any other trans-2-enoyl-CoA reductase classified under EC 1.3.1.38, EC 1.3.1.44, or EC 1.3.1.8; and/or   (d) said polypeptide having the activity of an esterase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 15 or 18 or is any other esterase classified under EC 3.1.1.1, EC 3.1.1.6, or EC 3.1.1.85.   
     
     
         7 .- 11 . (canceled) 
     
     
         12 . The method of  claim 1 , wherein 5-acetyloxypentanoyl-CoA is enzymatically synthesized from 5-hydroxypentanoyl-CoA using:
 (a) a polypeptide having the activity of an alcohol-O-acetyltransferase; or   (b) (i) a polypeptide having the activity of an alcohol-O-acetyltransferase and (ii) a polypeptide having the activity of a CoA transferase or a CoA lipase.   
     
     
         13 . (canceled) 
     
     
         14 . The method of  claim 12 , wherein said polypeptide having the activity of an alcohol-O-acetyltransferase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 17. 
     
     
         15 . A method for biosynthesizing 7-hydroxyheptanoate, said method comprising enzymatically synthesizing 7-acetyloxy-3-oxoheptanoyl-CoA from 5-acetyloxypentanoyl-CoA using a polypeptide having the activity of a β-ketothiolase or synthase classified under EC. 2.3.1.- and enzymatically converting 7-acetyloxy-3-oxoheptanoyl-CoA to 7-hydroxyheptanoate. 
     
     
         16 . The method of  claim 15 , wherein 7-acetyloxy-3-oxoheptanoyl-CoA is converted to 7-acetyloxy-3-hydroxyheptanoyl-CoA using:
 (a) a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, 7-acetyloxy-3-hydroxyheptanoyl-CoA is converted to 7-acetyloxy-hept-2-enoyl-CoA using a polypeptide having the activity of an enoyl-CoA hydratase, 7-acetyloxy-hept-2-enoyl-CoA is converted to 7-acetyloxyheptanoyl-CoA using a polypeptide having the activity of a trans-2-enoyl-CoA reductase, 7-acetyloxyheptanoyl-CoA is converted to 7-acetyloxyheptanoic acid using a polypeptide having the activity of a thioesterase or a CoA transferase, and 7-acetyloxyheptanoic acid is converted to 7-hydroxyheptanoic acid using a polypeptide having the activity of an esterase; or   (b) a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, 7-acetyloxy-3-hydroxyheptanoyl-CoA is converted to 7-acetyloxy-hept-2-enoyl-CoA using a polypeptide having the activity of an enoyl-CoA hydratase, 7-acetyloxy-hept-2-enoyl-CoA is converted to 7-acetyloxyheptanoyl-CoA using a polypeptide having the activity of a trans-2-enoyl-CoA reductase, 7-acetyloxyheptanoyl-CoA is converted to 7-hydroxyheptanoyl-CoA using a polypeptide having the activity of an esterase, and 7-hydroxyheptanoyl-CoA is converted to 7-hydroxyheptanoic acid using a polypeptide having the activity of a thioesterase or a CoA transferase.   
     
     
         17 . (canceled) 
     
     
         18 . The method of  claim 5  or  claim 15 , said method further comprising enzymatically converting 7-hydroxyheptanoate to pimelic acid, 7-aminoheptanoate, heptamethylenediamine, or 1,7 heptanediol in one or more steps. 
     
     
         19 . The method of  claim 18 , wherein:
 (a) 7-hydroxyheptanoate is converted to pimelic acid using one or more of a polypeptide having the activity of a monooxygenase, a primary alcohol dehydrogenase, a 7-hydroxyheptanoatedehydrogenase, a 7-oxoheptanoate dehydrogenase, a 6-oxohexanoate dehydrogenase, a 5-oxovalerate dehydrogenase, or an aldehyde dehydrogenase;   (b) 7-hydroxyheptanoate is converted to 7-aminoheptanoate using one or more of a polypeptide having the activity of a primary alcohol dehydrogenase, a 7-hydroxyheptanoatedehydrogenase, a 5-hydroxypentanoate dehydrogenase, a 4-hydroxybutyrate dehydrogenase, and a ω-transaminase;   (c) further comprising converting 7-aminoheptanoate to heptamethylenediamine using one or more of a polypeptide having the activity of a carboxylate reductase and a ω-transaminase;   (d) 7-hydroxyheptanoate is converted to heptamethylenediamine using one or more of a polypeptide having the activity of a carboxylate reductase, a ω-transaminase, a primary alcohol dehydrogenase, an N-acetyltransferase, and an acetylputrescine deacylase; and/or   (e) 7-hydroxyheptanoate is converted to 1,7 heptanediol using a polypeptide having the activity of a carboxylate reductase and an alcohol dehydrogenase.   
     
     
         20 .- 22 . (canceled) 
     
     
         23 . The method of  claim 19 , wherein:
 (a) said polypeptide having the activity of a ω-transaminase has at least 70% sequence identity to any one of the amino acid sequences set forth in SEQ ID NOs: 7-12; and/or   (b) said polypeptide having the activity of a carboxylate reductase has at least 70% sequence identity to any one of the amino acid sequences set forth in SEQ ID NOs 2-6 or 29.   
     
     
         24 .- 25 . (canceled) 
     
     
         26 . The method of  claim 1  or  claim 15 , wherein said 5-acetyloxypentanoyl-CoA is enzymatically produced from 2-oxoadipate. 
     
     
         27 . The method of  claim 26 , wherein 5-acetyloxypentanoyl-CoA is enzymatically produced from 2-oxoadipate using one or more of a polypeptide having the activity of a glutamate synthase, a 2-oxoglutarate decarboxylase, a branch chain decarboxylase, a glutamate decarboxylase, a ω-transaminase, a CoA transferase, a CoA ligase, an alcohol-O-acetyltransferase, and an alcohol dehydrogenase. 
     
     
         28 . The method of  claim 1  or  claim 15 , wherein said method is performed in a recombinant host. 
     
     
         29 . The method of  claim 28 , wherein:
 (a) said host is subjected to a cultivation strategy under aerobic, anaerobic or, micro-aerobic cultivation conditions;   (b) said host is cultured under conditions of nutrient limitation;   (c) said host is retained using a ceramic hollow fiber membrane;   (d) the principal carbon source fed to the fermentation derives from a biological feedstock; and/or   (e) the principal carbon source fed to the fermentation derives from a non-biological feedstock.   
     
     
         30 .- 32 . (canceled) 
     
     
         33 . The method of  claim 29 , wherein the biological feedstock is, or derives from, monosaccharides, disaccharides, lignocellulose, hemicellulose, cellulose, lignin, levulinic acid, formic acid, triglycerides, glycerol, fatty acids, agricultural waste, condensed distillers' solubles, or municipal waste. 
     
     
         34 . (canceled) 
     
     
         35 . The method of  claim 29 , wherein the non-biological feedstock is, or derives from, natural gas, syngas, CO 2 /H 2 , methanol, ethanol, benzoate, non-volatile residue (NVR) caustic wash waste stream from cyclohexane oxidation processes, or terephthalic acid/isophthalic acid mixture waste streams. 
     
     
         36 . The method of  claim 28 , wherein the host is a prokaryote or a eukaryote. 
     
     
         37 . The method of  claim 36 , wherein said prokaryote is from a genus selected from the group consisting of  Escherichia, Clostridia, Corynebacteria, Cupriavidus, Pseudomonas, Delftia, Bacillus, Lactobacillus, Lactococcus  and  Rhodococcus.    
     
     
         38 . The method of  claim 37 , wherein said prokaryote is selected from the group consisting of  Escherichia coli, Clostridium ljungdahlii, Clostridium autoethanogenum, Clostridium kluyveri, Corynebacterium glutamicum, Cupriavidus necator, Cupriavidus metallidurans, Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas oleavorans, Delftia acidovorans, Bacillus subtilis, Lactobacillus delbrueckii, Lactococcus lactis , and  Rhodococcus equi.    
     
     
         39 . (canceled) 
     
     
         40 . The method of  claim 36 , wherein said eukaryote is from a genus selected from the group consisting of  Aspergillus, Saccharomyces, Pichia, Yarrowia, Issatchenkia, Debaryomyces, Arxula , and  Kluyveromyces.    
     
     
         41 . The method of  claim 40 , wherein said eukaryote is selected from the group consisting of  Aspergillus niger, Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, Issathenkia orientalis, Debaryomyces hansenii, Arxula adenoinivorans , and  Kluyveromyces lactis.    
     
     
         42 . The method of  claim 28  wherein:
 (a) the host's tolerance to high concentrations of a C7 building block is improved through continuous cultivation in a selective environment; 
 (b) said host comprises an attenuation of one or more of the following enzymes: a polyhydroxyalkanoate synthase, an acetyl-CoA thioesterase, a phosphotransacetylase forming acetate, an acetate kinase, a lactate dehydrogenase, a menaquinol-fumarate oxidoreductase, an alcohol dehydrogenase forming ethanol, a triose phosphate isomerase, a pyruvate decarboxylase, a glucose-6-phosphate isomerase, an NADH-consuming transhydrogenase, an NADH-specific glutamate dehydrogenase, an NADH/NADPH-utilizing glutamate dehydrogenase, a pimeloyl-CoA dehydrogenase, an acyl-CoA dehydrogenase accepting C6 building blocks and central precursors as substrates, a butyryl-CoA dehydrogenase, or an adipyl-CoA synthetase; and/or 
 (c) said host overexpresses one or more genes encoding: an acetyl-CoA synthetase, a 6-phosphogluconate dehydrogenase, a transketolase, a puridine nucleotide transhydrogenase, a glyceraldehyde-3P-dehydrogenase, a malic enzyme, a glucose-6-phosphate dehydrogenase, a glucose dehydrogenase, a fructose 1,6 diphosphatase, a L-alanine dehydrogenase, a L-glutamate dehydrogenase, a formate dehydrogenase, a L-glutamine synthetase, a diamine transporter, a dicarboxylate transporter, and/or a multidrug transporter. 
 
     
     
         43 .- 44 . (canceled) 
     
     
         45 . A non-naturally occurring recombinant host comprising at least one exogenous nucleic acid encoding (i) an alcohol O-acetyltransferase, (ii) a β-ketothiolase or synthase, (iii) a thioesterase or a CoA transferase, (v) an esterase and one or more of (vi) a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, (vii) an enoyl-CoA hydratase, and (viii) a trans-2-enoyl-CoA reductase, said host producing 7-hydroxyheptanoate. 
     
     
         46 . The recombinant host of  claim 45 , wherein:
 (a) said host further comprising one or more of the following exogenous enzymes: a monooxygenase, an alcohol dehydrogenase, a 5-oxovalerate dehydrogenase, a 7-hydroxyheptanoatedehydrogenase, a 7-oxoheptanoate dehydrogenase, a 6-oxohexanoate dehydrogenase, or an aldehyde dehydrogenase, said host further producing pimelic acid;   (b) said host further comprising one or more of the following exogenous enzymes: a transaminase, a 7-hydroxyheptanoatedehydrogenase, a 5-hydroxypentanoate dehydrogenase, a 4-hydroxybutyrate dehydrogenase, and a primary alcohol dehydrogenase, said host further producing 7-aminoheptanoate;   (c) said host further comprising one or more of the following exogenous enzymes: a carboxylate reductase, a ω-transaminase, a deacylase, an N-acetyl transferase, or a primary alcohol dehydrogenase, said host further producing heptamethylenediamine; and/or   (d) said host further comprising an exogenous carboxylate reductase and an exogenous primary alcohol dehydrogenase, said host further producing 1,7 heptanediol.   
     
     
         47 .- 49 . (canceled) 
     
     
         50 . The recombinant host of  claim 45 , said host further comprising one or more of the following exogenous enzymes: a glutamate synthase, a 2-oxoglutarate decarboxylase, a branch-chain decarboxylase, a glutamate decarboxylase, a ω-transaminase, a CoA-ligase, a CoA-transferase, and an alcohol dehydrogenase. 
     
     
         51 . The recombinant host of  claim 45 , wherein:
 (a) said β-ketothiolase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1 or SEQ ID NO: 16;   (b) said enoyl-CoA reductase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 23-28;   (c) said alcohol O-acetyltransferase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 17;   (d) said carboxylate reductase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 2-6 or 29;   (e) said ω-transaminase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NOs: 7-12; and/or   (f) said esterase has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 15 or SEQ ID NO: 18.   
     
     
         52 .- 56 . (canceled) 
     
     
         57 . A biochemical network comprising a β-ketothiolase or synthase classified under EC. 2.3.1.-, 5-acetyloxypentanoyl-CoA, and 7-acetyloxy-3-oxoheptanoyl-CoA, wherein said β-ketothiolase or synthase enzymatically converts 5-acetyloxypentanoyl-CoA to 7-acetyloxy-3-oxoheptanoyl-CoA. 
     
     
         58 . The biochemical network of  claim 57 , further comprising a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, an enoyl-CoA hydratase, a trans-2-enoyl-CoA reductase, an esterase, and a thioesterase or a CoA transferase, wherein said 3-hydroxyacyl-CoA dehydrogenase or said 3-oxoacyl-CoA reductase, said enoyl-CoA hydratase, said trans-2-enoyl-CoA reductase, said esterase, and said thioesterase or said CoA transferase enzymatically convert 7-acetyloxy-3-oxoheptanoyl-CoA to 7-hydroxyheptanoate. 
     
     
         59 . A means for producing 7-acetyloxy-3-oxoheptanoyl-CoA, wherein said means enzymatically converts 5-acetyloxypentanoyl-CoA to 7-acetyloxy-3-oxoheptanoyl-CoA. 
     
     
         60 . The means of  claim 59 , wherein said means comprises a β-ketothiolase or synthase classified under EC. 2.3.1.-. 
     
     
         61 . The means of  claim 59 , further comprising means for enzymatically converting 7-acetyloxy-3-oxoheptanoyl-CoA to 7-hydroxyheptanoate. 
     
     
         62 . The means of  claim 61 , wherein said means comprises a 3-hydroxyacyl-CoA dehydrogenase or a 3-oxoacyl-CoA reductase, an enoyl-CoA hydratase, a trans-2-enoyl-CoA reductase, an esterase, and a thioesterase or a CoA transferase. 
     
     
         63 . A step for obtaining 7-acetyloxy-3-oxoheptanoyl-CoA using a β-ketothiolase or synthase classified under EC. 2.3.1.-. 
     
     
         64 . A composition comprising a bio-based, bio-derived or fermentation-derived 5-acetyloxypentanoyl-CoA, bio-based, bio-derived or fermentation-derived 7-acetyloxy-3-oxoheptanoyl-CoA, and a β-ketothiolase or synthase classified under EC. 2.3.1.-. 
     
     
         65 . A composition comprising bio-based, bio-derived or fermentation-derived 7-acetyloxy-3-oxoheptanoyl-CoA. 
     
     
         66 . The composition of  claim 64  or  claim 65 , wherein said composition is acellular or cellular. 
     
     
         67 . (canceled) 
     
     
         68 . A bio-based, bio-derived or fermentation-derived 7-acetyloxy-3-oxoheptanoyl-CoA produced by the method of enzymatically converting 5-hydroxypentanoate to 7-acetyloxy-3-oxoheptanoyl-CoA in one or more enzymatic steps using (i) a β-ketothiolase or synthase and (ii) and an alcohol O-acetyltransferase classified under EC. 2.3.1.-. 
     
     
         69 . A bio-derived product, bio-based product or fermentation-derived product, wherein said product comprises:
 i. a composition comprising at least one bio-derived, bio-based or fermentation-derived compound produced according to  claim 1  or  claim 15 , or any one of  FIGS. 1-5 , or any combination thereof,   ii. a bio-derived, bio-based or fermentation-derived polymer comprising the bio-derived, bio-based or fermentation-derived composition or compound of i., or any combination thereof,   iii. a bio-derived, bio-based or fermentation-derived resin comprising the bio-derived, bio-based or fermentation-derived compound or bio-derived, bio-based or fermentation-derived composition of i. or any combination thereof or the bio-derived, bio-based or fermentation-derived polymer of ii. or any combination thereof,   iv. a molded substance obtained by molding the bio-derived, bio-based or fermentation-derived polymer of ii. or the bio-derived, bio-based or fermentation-derived resin of iii., or any combination thereof,   v. a bio-derived, bio-based or fermentation-derived formulation comprising the bio-derived, bio-based or fermentation-derived composition of i., bio-derived, bio-based or fermentation-derived compound of i., bio-derived, bio-based or fermentation-derived polymer of ii., bio-derived, bio-based or fermentation-derived resin of iii., or bio-derived, bio-based or fermentation-derived molded substance of iv, or any combination thereof, or   vi. a bio-derived, bio-based or fermentation-derived semi-solid or a non-semi-solid stream, comprising the bio-derived, bio-based or fermentation-derived composition of i., bio-derived, bio-based or fermentation-derived compound of i., bio-derived, bio-based or fermentation-derived polymer of ii., bio-derived, bio-based or fermentation-derived resin of iii., bio-derived, bio-based or fermentation-derived formulation of v., or bio-derived, bio-based or fermentation-derived molded substance of iv., or any combination thereof.   
     
     
         70 . A non-naturally occurring organism comprising at least one exogenous nucleic acid encoding at least one polypeptide having the activity of at least one enzyme depicted in any one of  FIGS. 1 to 5 . 
     
     
         71 . A nucleic acid construct or expression vector comprising
 (a) a polynucleotide encoding a polypeptide having the activity of a β-ketothiolase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a β-ketothiolase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 1 or SEQ ID NO: 16;   (b) a polynucleotide encoding a polypeptide having the activity of a trans-2-enoyl-CoA reductase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a trans-2-enoyl-CoA reductase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27 or SEQ ID NO: 28;   (c) a polynucleotide encoding a polypeptide having the activity of ω-transaminase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of ω-transaminase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, or SEQ ID NO: 12;   (d) a polynucleotide encoding a polypeptide having the activity of a phosphopantetheinyl transferase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having phosphopantetheinyl transferase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 13 or 14;   (e) a polynucleotide encoding a polypeptide having the activity of an alcohol-O-acetyltransferase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of an alcohol-O-acetyltransferase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 17;   (f) a polynucleotide encoding a polypeptide having the activity of a carboxylate reductase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a carboxylate reductase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6 or SEQ ID NO: 29;   (g) a polynucleotide encoding a polypeptide having the activity of a esterase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a esterase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 15;   (h) a polynucleotide encoding a polypeptide having the activity of a pimeloyl-[acp] methyl ester esterase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a pimeloyl-[acp] methyl ester esterase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 18;   (i) a polynucleotide encoding a polypeptide having the activity of a CoA-transferase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a CoA-transferase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 19; or   (j) a polynucleotide encoding a polypeptide having the activity of a decarboxylase, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct production of the polypeptide and wherein the polypeptide having the activity of a decarboxylase is selected from the group consisting of a polypeptide having at least 70% sequence identity to the polypeptide of SEQ ID NO: 20, SEQ ID NO: 21, or SEQ ID NO: 22.   
     
     
         72 . A composition comprising the nucleic acid construct or expression vector of  claim 71 .